278 General Notes. 
Van Beneden (Nouvelles Recherches sur la fécondation, ete., 1887) 
has employed a stronger mixture, consisting of absolute alcohol and 
acetic acid in equal parts, without the addition of osmic acid. 
Preparation of Material—1. Freshly obtained! Ascaris females 
are placed between two sheets of cotton, which have been moistened 
a little in a 3 per cent. solution of common salt, then covered with 
a bell glass, and exposed one to three hours to an incubation tem- 
perature of 25°C. This procedure brings the polar globules to 
development in the younger eggs, and forces the cleavage in the 
older eggs. 
2. After an hour’s incubation, it is well to preserve a part of the 
material at disposal. The genital sacks are laid bare by a longitu- 
dinal slit in the body-wall, opposite the sexual aperture; the vagina 
is then cut free from the body, the alimentary tract lying between 
the two sacks is carefully removed, and the ovarian portions of the 
sacks are cut off, leaving the uterine portions with their contents 
for preservation. The anterior ends of the uteri contain eggs in 
all stages of maturation and fecundation ; the posterior ends contain 
eggs already beginning to cleave. The killing and hardening pro- 
cess should vary considerably for these different stages. 
3. It is advisable, therefore, to cut each uterus into thirds, and 
to expose the anterior third to the action of the acid mixture only 
five to seven minutes, the middle third ten to fifteen minutes, and 
the posterior third at least twenty-five minutes. After fixation, the 
anterior and middle thirds are transferred to 30 per cent. alcohol, 
and after a few hours to 50 per cent. alcohol, in which they may be 
kept for a long time. Eggs in process of cleavage—found in the 
posterior third—should be removed to absolute alcohol the moment 
they begin to show a light brown staining. After two to three 
hours they are to be transferred to 70 per cent. alcohol for preser- 
vation. If the acid mixture be heated to about 24°C., the poste- 
rior third of the uterus will require an exposure of only ten to fif- 
teen minutes. : 
4. Schneider’s acid carmine is an excellent staining agent. It 1s 
prepared as follows: Glacial acetic acid is diluted with distilled 
water to about 50 per cent.; then as much pulverized carmine 1s 
added to the boiling acid as will dissolve. - After filtering until the 
fluid becomes clear, a little rectified wood-vinegar is added (one 
drop A. pyrolignosum to ten ccm. of the carmine solution) for the 
purpose of strengthening the clarifying power of the mixture. 
he younger stages may be left in the dye three to four hours, 
the older stages eight to ten hours. 
Beautiful views of the karyokinetic figures are thus obtained, but 
they are not permanent. After three to four hours they begin to 
lose in distinctness. 
1 From the living horse, by means of arsenic pills. 
